In adult dogs after pneumonectomy (PNX), mechanical forces on the remaining lung are thought to stimulate compensatory lung growth. Following moderate (42% of total) resection, lobar growth is non-uniform. As resection increases (58-70%), alveolar growth is stimulated in all remaining lobes probably due to mechanical forces exceeding a threshold for cellular stimulation. Supplementation with all trans-retinoic acid (RA), a promoter of cell growth and differentiation, enhanced some aspects of alveolar-capillary response following 58% but not 42% resection, suggesting a relationship between the intensity of endogenous stimuli and the capacity for pharmacological growth enhancement. Lobar expansion explains ~40-70% of post-PNX compensation, presumably via increased septal strain/shear;residual compensation might be explained by elevated lobar perfusion via increased strain/shear on alveolar capillaries. Lobar expansion may also alter and interact with regional perfusion. Relationship of in vivo tissue mechanical stress to cellular response has not been established, but the hypoxia-inducible factor-erythropoietin-vascular endothelial growth factor (HIF-EPO- VEGF) axis is implicated. Our hypotheses are: 1) Regional compensatory lung growth varies directly with the intensity of regional mechanical stimuli, and 2) Growth factor supplementation amplifies natural response to post-PNX mechanical stimuli, but cannot initiate de novo compensatory lung growth in the absence of sufficient endogenous stimuli. In Aim 1, we will quantify post-PNX regional lung strain and shear using bronchovascular landmarks imaged at different transpulmonary pressures by high resolution computed tomography (HRCT) and correlate strain/shear with regional cellular response in HIF-EPO-VEGF axis. In Aim 2, we will test the effects of altering lobar perfusion on post-PNX compensation by banding one lobar pulmonary artery, which restrict its perfusion while exaggerating post-PNX perfusion to unbanded lobes. Regional perfusion will be measured by fluorescent microspheres. The remaining lobes will be assessed for cell signaling and for alveolar ultrastructure by morphometry. In Aim 3, we will establish cause-effect mechanisms of mechanical stresses on growth- related signaling in vitro using cultured lung epithelial cells subjected to mechanical strain and endothelial cells subjected to strain and fluid shear. Mechano-stress response of HIF-EPO-VEGF axis will be compared to its O2-sensitive response. In vitro signaling events will be related to parallel in vivo stress response. Epithelial- endothelial co-cultures will test for diffusible mediators of mechanotransduction. In Aim 4, we will amplify EPO/EPO-R signaling post-PNX by nebulization of recombinant human erythropoietin (rhEPO) to determine its local effect on angiogenesis, growth and function in relation to in vivo regional mechanical stresses. These studies define fundamental mechanical stimuli-response relationships in a robust model of compensatory lung growth, and explore a potential intervention for augmenting the endogenous response. Results have important basic and clinical implications for the re-initiation of lung growth in human chronic lung disease. PUBLIC HEALTH RELEVANCE: When one lung is surgically removed, the remaining lung undergoes compensatory growth where more lung tissue is generated to improve lung function. Our goal is to understand the stimuli and regulatory pathways responsible for re-initiating growth in the adult lung, specifically those pathways related to mechanical forces that stretch and distort the remaining tissue and blood vessels. We will study the intact lung as well as lung cells grown in culture subjected to mechanical distortion. These studies will advance fundamental knowledge of lung regeneration, and explore a potential intervention for augmenting the natural response. The results have important implications for the stimulation of lung re-growth in the treatment of human lung disease.